The vascularization of engineered tissues is critical to the ultimate success of tissue engineering as an organ replacement therapy. The formation of new capillary vessels in vivo, or angiogenesis, is linked to the pathogenesis of numerous diseases including cancer, and is regulated by local cues within the tissue microenvironment. The general goal of this RENEWAL proposal is to understand the mechanism by which local extracellular matrix (ECM) properties regulate capillary endothelial cell proliferation and capillary tube morphogenesis required in angiogenesis. The investigator has found that adhesion to ECM cooperates with growth factors to generate not only biochemical, but also mechanical signals that are important to driving capillary endothelial cell function. Studies from the past grant period demonstrated that the spatial organization and geometric presentation of ECM affects the degree to which cells attach and spread against the substrate. These changes in cell shape regulated proliferation and capillary tube formation by modulating contractile tension generated by the actin cytoskeleton. It was also found that changes in the mechanical stiffness of the ECM also regulated proliferation and tubulogenesis, likely by modulating similar mechanical signals. In this proposal, the investigator proposes to further investigate the role of these structural and mechanical cues in regulating angiogenic behaviors. Specific Aim 1 will be to investigate the cooperation between ECM geometry, soluble factors, and cell mechanics in regulating capillary growth and differentiation. Specific Aim 2 will be to investigate the role of ECM stiffness in regulating cell shape, mechanics, and EC function. Specific Aim 3 will be to examine the role of mechanical cues in regulating angiogenic responses in 3-dimensional tissue microenvironments. Together, these studies will define the mechanisms by which local structural and mechanical properties within ECM modulate cell function and capillary morphogenesis, and establish new strategies to promote angiogenesis in native ischemic tissues as well as in ex-vivo engineered tissues.